Reversible redox post-translational modifications on protein thiols have been implicated in several signaling pathways of biological importance. Protein S-nitrosylation and proteins S-sulfinylation are two of these modifications that play critical roles in maintaining the redox balance of proteins. Redox imbalance has recently been shown to play a crucial role in heart disease, neurodegeneration and cancer. Protein S-nitrosylation describes the reversible, post-translational modification of select thiols with nitric oxide (NO) and/or its oxidized products to form S-nitrosothiols (SNO). Protein S-sulfinylation describes the oxidation of cysteine thiols to a SOOH (sulfinic acid) motif via a peroxide-mediated pathway.
Current methods to study S-nitrosylation and S-sulfinylation lack sufficient selectivity, and may over-represent the functional role of S-nitrosylation and S-sulfinylation.
For example, several methods have been reported to label and enrich sites of protein S-nitrosylation, including several versions of the biotin switch technique (BST), gold nanoparticle based enrichment, organomercury based methods and phosphine-based probes. By far, the most popular method is BST, which relies on ascorbate reduction to selectively reduce sites of S-nitrosylation. Sodium ascorbate has been shown to reduce activated disulfides and its reactivity with cysteine sulfenic acids and thiocysteines has not been thoroughly investigated. This suggests that the BST may be contaminated with weak disulfides, thiosulfhydrylation (R-SSH), or other activated thiol modifications. Such findings demonstrate that knowledge about SNO modifications derived from such methods are indirect.
New selective methods for detecting S-nitrosylation and S-sulfinylation are needed.